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The reaction flask was sealed, covered by aluminum foil to prevent the light, taken out from the glovebox, and stirred at 40 C in an oil bath for 24 h

This buffer accounts for normal losses and provides flexibility if you decide to extend your protocol or adjust dosing

What results should you expect

Specifically, p66Shc can generate ROS through the oxidation of cytochrome c by utilizing the reducing equivalents of the mitochondrial ETC, playing a crucial role in carcinogenesis and aging [44]

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If this happens, simply lowering your dose usually solves the problem